cDNA and protein characterization of human MAGE-10

Abstract

ADAGE genes are frequently expressed in several types of human malignancy and code for antigens recognized by cytotoxic T lymphocytes. We have previously described a monoclonal antibody (MAb), named 6CI, that recognizes the MAGE-I protein and cross-reacts with a 72-kDa protein present in lysates of melanoma cells such as MZ2-MEL. To identify this protein, we have screened an expression library prepared from MZ2-MEL cells. Several clones that encoded a protein recognized by antibody 6CI contained a sequence identical to that of MAGE-10, another member of the MAGE-A gene family. Full- length MAGE-10 cDNA clones, obtained after screening additional cDNA melanoma libraries, were found to be approximately 2.5 kb in length. In vitro translation and transient transfection experiments indicated that MAGE-10 codes for a protein of approximately 72 kDa. This product was recognized by MAb 6CI as well as by a polyclonal serum raised against a MAGE-10 peptide, thus demonstrating its identity with MAGE-10. Analysis of MAGE-10 mRNA by RT- PCR confirmed its presence in testis and placenta but not in other normal tissues. Expression-of MAGE-10 in melanoma tumors was found to parallel that of MAGE-I. Western blot analysis with the polyclonal anti-MAGE-10 antibody showed the presence of MAGE-10 in lysates of purified trophoblast cells. Immuno-cytochemistry of cultured melanoma cells indicated that MAGE-10 is a nuclear protein.